Studies on the non-linear osmotic pressure-volume relationship in mitochondria and entry of sucrose into the matrix space during centrifugation

Abstract

The two-compartment sucrose-space hypothesis was refuted recently (Sitaramam, V. and Sarma, M.K.J. (1981) Proc. Natl. Acad. Sci. U.S.A. 78, 3441-3445), using the novel technique of enzyme osmometry of matrix enzymes based on the following premise: a shift in the discontinuity (break-point) of the activity profile of an occluded enzyme as a function of external osmolarity (i.e., osmotic profiles) would imply a shift in the internal solute content of the same compartment as that of the enzyme. A systematic re-evaluation of the osmotic profiles of mitochondrial enzyme systems has revealed that the activities of several matrix and inner membrane enzyme systems exhibited break-points larger than those of osmolysis (i.e., actual release of marker enzymes into the medium) of mitochondria. The experimental findings were consistent with (i) entry of sucrose across the inner membrane consequent to enhanced permeability effects by gravitational field, and (ii) dependence of the kinetic constant of several membrane-bound enzymes/carrier proteins, respiratory coupling and ADP/O ratio on the volume of mitochondria (i.e., osmotic stretch of the inner membrane).